An ovoperoxidase is present in the cortical granules of mouse oocytes. Following cortical reaction ovoperoxidase is detectable either in the perivitelline space or on the surface of the zona pellucida. The enzyme is not detectable in the presence of peroxidase inhibitors. Results indicate that the ovoperoxidase is apparently responsible for the hardening reaction of the zona pellucida following cortical reaction. This conclusion is based on the following observations: (1) both ovoperoxidase activity and zona hardening are blocked by several known peroxidase inhibitors; (2) tyrosine analogs, which should competitively inhibit peroxidase catalyzed cross-linking of protein-bound tyrosines, also prevent hardening; and (3) horseradish peroxidase partially hardens the zona of unactivated eggs.